GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise based on Thalamus big data

GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise based on Thalamus big data | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Short Report GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise based on Thalamus big data Lin Wang, tao hong, Wenbin Cui, Minghu Jiang This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-2906322/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract G protein-coupled receptor (GPR68) is widely distributed in the human body and participates in various physiological and pathological processes. The main GPR68-inhibited and -activated molecular network was constructed in Thalamus from GDS2678 of NCBI by integrating SAM, SPSS Pearson and GRNInfer mutual positive verification based on richness. The main GPR68 inhibited molecular subnetwork composed of upstream ALG8, or MAF, feedback SMA4, and downstream GSTM3_1 with TYMS. GPR68 activated molecular subnetwork contained upstream RAD50, feedback SPAG9, and downstream RFK. The relationship of SMA4 inhibition to SPAG9 -> MAF -> RAD50 -> ALG8 was found in Thalamus for negative main molecular verification. The relationship of ALG8 activation to SMA4 -> GSTM3_1 -> TYMS -> MAF in Thalamus for positive main molecular verification. Our results show GPR68 inhibited SMA4-nucleoplasm RNA splicing coupling ALG8, or MAF-nucleus sequence specific DNA binding, or ER amino acid glycosylation to GSTM3_1 with TYMS-extracellular exosome small molecule-based circadian exercise, and GPR68 activated SPAG9-integral component of membrane kinesin binding coupling RAD50-membrane positive regulation of kinase activity to RFK-mitochondrion apoptotic process for mutual negative knowledge verification. We put forwards GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise based on Thalamus big data. Our model is positive verified by the other similar and common inhibited and/or activated knowledge in Thalamus. The role and mechanism of our GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise contributes to better detection, evaluation, intervention, tracking, and other health management of brain diseases. GPR68 inhibited Thalamus subnetwork GPR68 activated Thalamus subnetwork GPR68 inhibited nucleus or ER to exosome GPR68 inhibited circadian exercise GPR68 activated protein Figures Figure 1 Figure 2 Introduction GPR68 is widely distributed in the human body and participates in various physiological and pathological processes. Such as, GPR68 is also expressed in the nervous system, participating in a variety of physiological and pathological processes [ 1 – 4 ]. The abnormal expression of GPR68 is related to the occurrence and development of multiple cancer [ 5 – 13 ]. The current research mainly focuses on its regulatory mechanism[ 14 – 18 ], signaling pathways[ 19 – 23 ] and potential clinical applications. GPR68 locates plasma membrane, integral component of plasma membrane, functions as protein coupled receptor activity, inflammatory response, G protein coupled receptor signaling pathway, positive regulation of insulin secretion involved in cellular response to glucose stimulus, negative regulation of monocyte differentiation, cellular response to pH, positive regulation of osteoclast development, GPCRDB class A rhodopsin like, rhodopsin like receptor activity, osteoclasts by the Database for Annotation, Visualization and Integrated Discovery (DAVID) [ 24 , 25 ]. However, GPR68-inhibited nucleus or ER to exosome small molecule-based circadian exercise mechanism is not clear in Thalamus. We have already published several papers[ 26 – 29 ] from GDS2678[ 30 ] by integrating microarray significance analysis (SAM) [ 31 ], SPSS correlation coefficient Pearson and gene (protein) reconstruction network (GRNInfer) [ 32 ] mutual positive verification. GPR68-inhibited subnetwork we identified includes upstream ALG8, MAF, feedback SMA4, downstream GSTM3_1, GSTM3_2, NUP214, TYMS, UBB. GPR68-activated subnetwork includes upstream PER2, RAD50, feedback SPAG9, downstream CDR2, CGRRF1, DZIP3, ISCA1, RFK in GDS2678 in Thalamus. The main GPR68-inhibited and -activated molecular network will be constructed up/downstream/feedback in Thalamus for inhibited nucleus or ER to exosome non-protein based circadian exercise based on richness. The inhibition relationship in Thalamus for negative main molecular verification. The activation relationship will be found in Thalamus for positive main molecular verification. The main knowledge of GPR68-inhibited and -activated subnetwork will be constructed up/downstream/feedback for inhibited nucleus or ER to exosome non-protein based circadian exercise. We will integrate GPR68-inhibited and -activated knowledge and put forwards Thalamus model. GPR68-inhibited and -activated knowledge subnetwork will be demonstrated in Thalamus for mutual negative knowledge verification. and will be positive verified by the other similar and common inhibited and/or activated knowledge. The role and mechanism of our GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise contributes to better detection, evaluation, intervention, tracking, and other health management of brain diseases. Method 441 significant high expression molecules in 14 human left hemispheres were selected from 12,558 genes compared with the corresponding low expression of 15 chimpanzee left hemispheres in GDS2678 (public free from NCBI) by significance analysis of microarrays (SAM) ( http://www-stat.stanford.edu/~tibs/SAM/ ), Data were processed using a log base of two and two unpaired classes with minimum fold change (≥ 2). A false-discovery rate of 0% was chosen. GPR68-inhibited molecules in Thalamus was identified negative correlation coefficient (CC ≤ -0.25), including ALG8, MAF, SMA4, GSTM3_1, GSTM3_2, NUP214, TYMS, UBB, which are mutual positive Pearson correlation (CC ≥ 0.25) molecules except GPR68 by SPSS. GPR68 activated molecules from high expression Pearson mutual positive correlation coefficient (CC ≥ 0.25) were selected including PER2, RAD50, SPAG9, CDR2, CGRRF1, DZIP3, ISCA1, RFK in Thalamus by SPSS. GRNInfer is a tool used to construct all molecular activation and inhibition direction total network by linear programming and decomposition procedure defined by the following equation: $$\mathcal{J}=\left(\dot{\text{X}}-\text{B}\right)\text{U}{\text{E}}^{-1}{\text{V}}^{\text{T}}+\text{Y}{\text{V}}^{\text{T}}=\widehat{\mathcal{I}}+\text{Y}{\text{V}}^{\text{T}}$$ 1 The main GPR68-inhibited and -activated molecular network was constructed up/downstream/feedback in Thalamus for inhibited nucleus or ER to exosome non-protein based circadian exercise by GRNInfer based on richness. The inhibition relationship was found in Thalamus for negative main molecular verification. The activation relationship in Thalamus for positive main molecular verification. The main knowledge of GPR68-inhibited and activated subnetwork was constructed up/downstream/feedback for inhibited nucleus or ER to exosome non-protein based circadian exercise in Thalamus by DAVID, respectively, including GOTERM-BP-DIRECT, GOTERM-MF-DIRECT, KEGG-PATHWAY, BIOCARTA, GenMAPP, GOTERM-CC-DIRECT, GNF-U133A-QUARTILE UNIGENE-EST-QUARTILE. We integrated GPR68-inhibited and -activated knowledge and put forwards our model. GPR68-inhibited and -activated knowledge subnetwork was setup in Thalamus for mutual negative knowledge verification. Our model was positive verified by the corresponding other similar and common inhibited and/or activated knowledge. Results GPR68-inhibited and -activated network was constructed in Thalamus. GPR68 inhibited molecular subnetwork composed of upstream ALG8, MAF, feedback SMA4, and downstream GSTM3_1, GSTM3_2, NUP214, TYMS, UBB in Thalamus (Figure 1). GPR68 activated subnetwork included upstream PER2, RAD50, feedback SPAG9, and downstream CDR2, CGRRF1, DZIP3, ISCA1, RFK in Thalamus (Figure 1), based on mutual positive verification using CC and GRNInfer. The main GPR68 inhibited molecular subnetwork composed of upstream ALG8, or MAF, feedback SMA4, and downstream GSTM3_1 with TYMS (Figure 2). GPR68 activated molecular subnetwork contained upstream RAD50, feedback SPAG9, and downstream RFK (Figure 2). The relationship of SMA4 inhibition to SPAG9 -> MAF -> RAD50 -> ALG8 was found in Thalamus The relationship of ALG8 activation to SMA4 -> GSTM3_1 -> TYMS -> MAF in Thalamus (Figure 2). The main GPR68 inhibited knowledge subnetwork was identified upstream nucleus sequence specific DNA binding, or ER amino acid glycosylation, feedback nucleoplasm RNA splicing, and downstream extracellular exosome small molecule-based circadian exercise in Thalamus by DAVID, as shown in Table 1. The main GPR68 activated subnetwork included upstream membrane positive regulation of kinase activity, feedback integral component of membrane kinesin binding, and downstream mitochondrion apoptotic process in Thalamus by DAVID，as shown in Table 2. The main GPR68 inhibited common knowledge appears nucleus appears in the subnetwork of GPR68 inhibited-upstream MAF and inhibited-downstream GSTM3_1, TYMS in Thalamus. The main GPR68 activated common knowledge appears metal ion binding appears in the subnetwork of GPR68 activated-upstream RAD50 and activated-downstream RFK; ATP binding in the subnetwork of GPR68 activated-upstream RAD50 and activated-downstream RFK in Thalamus. The main GPR68 inhibited and activated common knowledge appears small molecule metabolic process appears in the subnetwork of GPR68 inhibited-downstream GSTM3_1, TYMS and activated-downstream RFK; protein binding in the subnetwork of GPR68 inhibited-upstream MAF and activated-upstream RAD50 and activated-feedback SPAG9; nucleoplasm in the subnetwork of GPR68 inhibited-feedback SMA4 and inhibited-downstream TYMS and activated-upstream RAD50; mitochondrion in the subnetwork of GPR68 inhibited-downstream TYMS and activated-downstream RFK; Metabolic pathways in the subnetwork of GPR68 inhibited-upstream ALG8 and inhibited-downstream TYMS and activated-downstream RFK; integral component of membrane in the subnetwork of GPR68 inhibited-upstream ALG8 and activated-feedback SPAG9; extracellular exosome in the subnetwork of GPR68 inhibited-downstream GSTM3_1 and activated-feedback SPAG9; cytosol in the subnetwork of GPR68 inhibited-downstream GSTM3_1, TYMS and activated-feedback SPAG9 and activated-downstream RFK; cytoplasm in the subnetwork of GPR68 inhibited-upstream MAF and inhibited-downstream GSTM3_1, TYMS and activated-feedback SPAG9 and activated-downstream RFK in Thalamus, as shown in Table 1 and 2. Discussion GPR68-inhibited and -activated network was constructed in Thalamus. GPR68 inhibited molecular subnetwork composed of upstream ALG8, MAF, feedback SMA4, and downstream GSTM3_1, GSTM3_2, NUP214, TYMS, UBB in Thalamus (Figure 1). GPR68 activated subnetwork included upstream PER2, RAD50, feedback SPAG9, and downstream CDR2, CGRRF1, DZIP3, ISCA1, RFK in Thalamus (Figure 1), based on mutual positive verification using CC and GRNInfer. The main GPR68 activated molecular subnetwork composed of upstream ALG8, or MAF, feedback SMA4, and downstream GSTM3_1 with TYMS. GPR68 inhibited molecular subnetwork contained upstream RAD50, feedback SPAG9, and downstream RFK (Figure 2). The relationship of SMA4 inhibition to SPAG9 -> MAF -> RAD50 -> ALG8 was found in Thalamus for negative main molecular verification. The relationship of ALG8 activation to SMA4 -> GSTM3_1 -> TYMS -> MAF in Thalamus for positive main molecular verification (Figure 2). The main GPR68 inhibited upstream molecule was determined as ALG8, or MAF in Thalamus by GRNInfer (Figure 1-2). The corresponding main inhibited upstream knowledge subnetwork was identified as nucleus sequence specific DNA binding, or ER amino acid glycosylation based on DAVID. We proposed GPR68 inhibited upstream nucleus sequence specific DNA binding, or ER amino acid glycosylation in Thalamus. ALG8 refers to ALG8 alpha-1,3-glucosyltransferase, an enzyme in the endoplasmic reticulum membrane and an integral component of the membrane. It has alpha 1,3 mannosyltransferase activity and dolichyl pyrophosphate Man9GlcNAc2 alpha 1,3 glucosyltransferase activity. ALG8 is involved in various biological processes, including protein N-linked glycosylation, dolichol-linked oligosaccharide biosynthetic process, oligosaccharide lipid intermediate biosynthetic process, protein N-linked glycosylation via asparagine, post-translational protein modification, cellular protein metabolic process, and mannosylation. ALG8 is associated with N-glycan biosynthesis and metabolic pathways. It is also related to glycoprotein metabolism, protein amino acid glycosylation, glycoprotein biosynthesis, and biopolymer glycosylation. MAF (v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog) is a protein present in chromatin, nucleus, and cytoplasm. it possesses transcriptional activator activity and sequence-specific DNA binding and protein binding capabilities. MAF is involved in various biological processes, such as negative regulation of transcription from RNA polymerase II promoter, cytokine production, regulation of transcription from RNA polymerase II promoter, transcription from RNA polymerase II promoter, regulation of chondrocyte differentiation, positive regulation of transcription from RNA polymerase II promoter, cell development, inner ear development, and lens fiber cell differentiation. It is associated with transcriptional misregulation in cancer and inflammatory bowel disease (IBD). MAF participates in activating the Th2 cytokine genes expression and has effects on RNA polymerase II transcription factor activity was selected by DAVID (Table 1) as positive verification for the main Thalamus knowledge. The main GPR68 inhibited feedback molecule was determined as SMA4 in Thalamus by GRNInfer (Figure 1-2). The corresponding main inhibited feedback knowledge subnetwork was identified as nucleoplasm RNA splicing based on DAVID. We proposed GPR68 inhibited feedback nucleoplasm RNA splicing in Thalamus. SMA4 refers to the glucuronidase beta pseudogene (SMA4), which can be found in the nucleoplasm, protein complex assembly, nuclear mRNA splicing via spliceosome, RNA splicing, RNA splicing via transesterification reactions with bulged adenosine as nucleophile, RNA splicing via transesterification reactions, and mRNA processing was selected by DAVID (Table 1) as positive verification for the main Thalamus knowledge. The main GPR68 inhibited downstream molecule was determined as GSTM3_1 with TYMS in Thalamus by GRNInfer (Figure 1-2). The corresponding main inhibited downstream knowledge subnetwork was identified as extracellular exosome small molecule-based circadian exercise based on DAVID. We proposed GPR68 inhibited downstream extracellular exosome small molecule-based circadian exercise in Thalamus. GSTM3_1 refers to glutathione S-transferase mu 3 (brain) (GSTM3), which can be found in the nucleus, cytoplasm, cytosol, sperm fibrous sheath, and extracellular exosome. It has glutathione transferase activity, enzyme binding ability, identical protein binding ability, protein homodimerization activity, and glutathione binding ability. GSTM3_1 is involved in various biological processes, such as glutathione metabolic process, xenobiotic metabolic process, establishment of blood nerve barrier, nitrobenzene metabolic process, xenobiotic catabolic process, response to estrogen, small molecule metabolic process, cellular detoxification of nitrogen compounds, and glutathione derivative biosynthetic process. GSTM3_1 is related to glutathione metabolism, metabolism of xenobiotics by cytochrome P450, drug metabolism cytochrome P450, and chemical carcinogenesis. Additionally, it has an association with circadian exercise. TYMS (Thymidylate Synthetase) is an enzyme that is present in the nucleus, nucleoplasm, nucleolus, cytoplasm, mitochondrion, mitochondrial inner membrane, mitochondrial matrix, and cytosol. It possesses nucleotide binding, mRNA binding, thymidylate synthase activity, folic acid binding, drug binding, protein homodimerization activity, and cofactor binding. TYMS is involved in various biological processes, including G1/S transition of mitotic cell cycle, regulation of transcription involved in G1/S transition of mitotic cell cycle, mitotic cell cycle, nucleobase containing compound metabolic process, pyrimidine nucleobase metabolic process, dTMP biosynthetic process, dTTP biosynthetic process, aging, circadian rhythm, cell proliferation, deoxyribonucleoside monophosphate biosynthetic process, response to toxic substance, cell growth, immortalization of host cell by virus, uracil metabolic process, organ regeneration, methylation, response to progesterone, response to vitamin A, response to cytokine, tetrahydrofolate interconversion, response to drug, small molecule metabolic process, response to ethanol, dUMP metabolic process, pyrimidine nucleoside biosynthetic process, response to organophosphorus, developmental growth, cartilage development, response to glucocorticoid, response to folic acid, nucleobase containing small molecule metabolic process, intestinal epithelial cell maturation, and DNA biosynthetic process. TYMS is involved in metabolic pathways such as Pyrimidine metabolism and One carbon pool by folate was selected by DAVID (Table 1) as positive verification for the main Thalamus knowledge. There are literature supporting the relationship between GPR68 and RAD50 families [33-36]. The main GPR68 activated upstream molecule was determined as RAD50 in Thalamus by GRNInfer (Figure 1-2). The corresponding main activated upstream knowledge subnetwork was identified as membrane positive regulation of kinase activity based on DAVID. We proposed GPR68 activated upstream membrane positive regulation of kinase activity in Thalamus. RAD50 refers to RAD50 double-strand break repair protein (RAD50), which can be found in the nuclear chromosome telomeric region, nucleoplasm, membrane, Mre11 complex, site of double-strand break, and pronucleus. RAD50 has single-stranded DNA endodeoxyribonuclease activity, DNA binding ability, ATP-dependent DNA helicase activity, protein binding ability, ATP binding ability, 3’ 5’ exonuclease activity, protein binding bridging ability, and metal ion binding ability. RAD50 is involved in the regulation of mitotic recombination, telomere maintenance, double-strand break repair via homologous recombination, DNA repair, double-strand break repair, double-strand break repair via nonhomologous end joining, DNA recombination, cellular response to DNA damage stimulus, telomere maintenance via telomerase, reciprocal meiotic recombination, regulation of mitotic cell cycle, viral process, telomeric 3’ overhang formation, positive regulation of protein autophosphorylation, positive regulation of telomere maintenance, DNA duplex unwinding, positive regulation of kinase activity, double-strand break repair via synthesis-dependent strand annealing, chromosome organization involved in meiosis, nucleic acid phosphodiester bond hydrolysis, and negative regulation of telomere capping. RAD50 is related to homologous recombination and non-homologous end joining. It is also involved in the h_atmPathway: ATM Signaling Pathway, h_atrbrcaPathway: Role of BRCA1, BRCA2 and ATR in Cancer Susceptibility. Additionally, RAD50 is related to the cell cycle, response to an endogenous stimulus, M phase, and response to DNA damage stimulus was selected by DAVID (Table 2) as positive verification for the main Thalamus knowledge. The main GPR68 activated feedback molecule was determined as SPAG9 in Thalamus by GRNInfer (Figure 1-2). The corresponding main activated feedback knowledge subnetwork was identified as integral component of membrane kinesin binding based on DAVID. We proposed GPR68 activated feedback integral component of membrane kinesin binding in Thalamus. SPAG9, which stands for sperm associated antigen 9, is a protein that is associated with various cellular components such as acrosomal vesicles, cytoplasm, microtubule organizing centers, cytosol, integral components of membranes, perinuclear regions of cytoplasm, and extracellular exosomes. It exhibits multiple molecular functions such as MAP kinase scaffold activity, protein binding, JUN kinase binding, kinesin binding, receptor signaling complex scaffold activity, and mitogen activated protein kinase p38 binding. Its biological processes include the activation of JUN kinase activity, spermatogenesis, positive regulation of cell migration, retrograde transport endosome to Golgi, muscle cell differentiation, positive regulation of neuron differentiation, striated muscle cell differentiation, positive regulation of muscle cell differentiation, protein homooligomerization, and negative regulation of protein homodimerization activity was selected by DAVID (Table 2) as positive verification for the main Thalamus knowledge. The main GPR68 activated downstream molecule was determined as RFK in Thalamus by GRNInfer (Figure 1-2). The corresponding main activated downstream knowledge subnetwork was identified as mitochondrion apoptotic process based on DAVID. We proposed GPR68 activated downstream mitochondrion apoptotic process in Thalamus. Riboflavin kinase (RFK) is a protein that is located in the cytoplasm, mitochondria, and cytosol. It exhibits molecular functions such as ATP binding, riboflavin kinase activity, and metal ion binding. RFK is involved in vitamin metabolic processes, including water-soluble vitamin metabolic processes, riboflavin metabolic processes, and riboflavin biosynthetic processes. It also participates in apoptotic processes, FMN biosynthetic processes, phosphorylation, positive regulation of NAD§H oxidase activity, small molecule metabolic processes, and reactive oxygen species metabolic processes. The functions of RFK are usually associated with riboflavin metabolism and metabolic pathways. Its important molecular function is magnesium ion binding was selected by DAVID (Table 2) as positive verification for the main Thalamus knowledge. There are literature supporting the activated relationship between GPR68 and RFK families[37, 38]. We put forwards the model of GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise based on Thalamus big data in Thalamus. GPR68 inhibited nucleoplasm RNA splicing coupling nucleus sequence specific DNA binding, or ER amino acid glycosylation to extracellular exosome small molecule-based circadian exercise, GPR68 activated integral component of membrane kinesin binding coupling membrane positive regulation of kinase activity to mitochondrion apoptotic process for mutual negative knowledge verification. The main GPR68-inhibited model is positive verified by common knowledge，including nucleus appears in the subnetwork of GPR68 inhibited-upstream MAF and inhibited-downstream GSTM3_1, TYMS in Thalamus. The main GPR68-activated model is positive verified by the common knowledge, including metal ion binding appears in the subnetwork of GPR68 activated-upstream RAD50 and activated-downstream RFK; ATP binding in the subnetwork of GPR68 activated-upstream RAD50 and activated-downstream RFK in Thalamus. The main GPR68 inhibited and activated model was positive verified by common knowledge, containing small molecule metabolic process appears in the subnetwork of GPR68 inhibited-downstream GSTM3_1, TYMS and activated-downstream RFK; protein binding in the subnetwork of GPR68 inhibited-upstream MAF and activated-upstream RAD50 and activated-feedback SPAG9; nucleoplasm in the subnetwork of GPR68 inhibited-feedback SMA4 and inhibited-downstream TYMS and activated-upstream RAD50; mitochondrion in the subnetwork of GPR68 inhibited-downstream TYMS and activated-downstream RFK; Metabolic pathways in the subnetwork of GPR68 inhibited-upstream ALG8 and inhibited-downstream TYMS and activated-downstream RFK; integral component of membrane in the subnetwork of GPR68 inhibited-upstream ALG8 and activated-feedback SPAG9; extracellular exosome in the subnetwork of GPR68 inhibited-downstream GSTM3_1 and activated-feedback SPAG9; cytosol in the subnetwork of GPR68 inhibited-downstream GSTM3_1, TYMS and activated-feedback SPAG9 and activated-downstream RFK; cytoplasm in the subnetwork of GPR68 inhibited-upstream MAF and inhibited-downstream GSTM3_1, TYMS and activated-feedback SPAG9 and activated-downstream RFK in Thalamus (Table 1 and 2). Summary The inhibition relationships among the main molecules were found in Thalamus for negative molecular verification. The activation relationships among the main molecules were found in Thalamus for positive molecular verification. We put forwards GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise in Thalamus, based on GPR68 inhibited nucleoplasm RNA splicing coupling nucleus sequence specific DNA binding, or ER amino acid glycosylation to extracellular exosome small molecule-based circadian exercise, and GPR68 activated integral component of membrane kinesin binding coupling membrane positive regulation of kinase activity to mitochondrion apoptotic process for mutual negative knowledge verification. Our model is positive verified by the other similar and common inhibited and/or activated knowledge in Thalamus. The role and mechanism of our GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise contributes to better detection, evaluation, intervention, tracking, and other health management of brain diseases. Declarations Ethics approval and consent to participate We have not Ethics because our data GSE10140-10141 public free from NCBI. Consent for publication All authors have approved the manuscript for submission. Data Availability Statement Our data GSE10140-10141 public free from NCBI. Competing interests We have not any competing interests with others. Funding This work is supported by grants from the National Natural Science Key Foundation of China (62036001), National Social Science Major Foundation of China (14ZDB154 * 15ZDB017), and the Independent scientific research project of Tsinghua University (20161080056). Author Contribution Statement LW not only has designed the whole experiment, put forward hypotheses and written the paper, but also established SAM and GRNInfer databases. TH has constructed molecular and knowledge network，Pearson positive correlation coefficient (CC) database. WBC has done programme and setup DAVID database. MHJ has done figures and tables and help analysis in this paper. All authors read and approved the final Manuscript. Acknowledgments Thank Zhenfu Jiang from Tencent for suggesting us many tools and methods. References Wang, T., et al., GPR68 Is a Neuroprotective Proton Receptor in Brain Ischemia. STROKE, 2020. 51(12): p. 3690–3700. Xu, Y., M.T. Lin and X. Zha, GPR68 deletion impairs hippocampal long-term potentiation and passive avoidance behavior (vol 13, pg 132, 2020). MOLECULAR BRAIN, 2021. 14(401). Xu, Y., M.T. Lin and X. Zha, GPR68 deletion impairs hippocampal long-term potentiation and passive avoidance behavior. MOLECULAR BRAIN, 2020. 13(1321). Nogueira, P.A.S., et al., The orphan receptor GPR68 is expressed in the hypothalamus and is involved in the regulation of feeding. NEUROSCIENCE LETTERS, 2022. 781(136660). Wiley, S.Z., et al., GPR68: An Emerging Drug Target in Cancer. INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES, 2019. 20(5593). Shore, D., et al., GPR68 limits the severity of chemical-induced oral epithelial dysplasia. Scientific reports, 2023. 13(1): p. 353–353. He, X., et al., Proton Sensor GPR68 Is Essential to Maintain Myeloid Malignancies. BLOOD, 2018. 1321. Elemam, N.M., et al., Expression Status of Proton Sensing Receptor GPR68 in Various Breast Cancer Molecular Subtypes. FASEB JOURNAL, 2022. 361. Elemam, N.M., et al., Expression of GPR68, an Acid-Sensing Orphan G Protein-Coupled Receptor, in Breast Cancer. FRONTIERS IN ONCOLOGY, 2022. 12(847543). Zhang, W., et al., Clinical data analysis reveals the role of OGR1 (GPR68) in head and neck squamous cancer. Animal models and experimental medicine, 2020. 3(1): p. 55–61. Wiley, S.Z., et al., GPR68, a proton-sensing GPCR that mediates interaction of pancreatic cancer associated fibroblasts and cancer cells, is a potential therapeutic target for pancreatic cancer. CANCER RESEARCH, 2019. 79S(13). Wiley, S., et al., GPR68, a proton sensing GPCR, mediates interaction of pancreatic cancer associated fibroblasts and cancer cells. FASEB JOURNAL, 2018. 32S(1). Wiley, S.Z., et al., GPR68, a proton-sensing GPCR, mediates interaction of cancer-associated fibroblasts and cancer cells. FASEB JOURNAL, 2018. 32(3): p. 1170–1183. Huang, X., et al., Allosteric ligands for the pharmacologically dark receptors GPR68 and GPR65. NATURE, 2015. 527(7579): p. 477-+. Bell, T.J., et al., Inhibition of Myofibroblast Differentiation and SMAD Mediated Transcription via Small Molecule Stimulation of GPR68. AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE, 2021. 203S(9). Yu, X., et al., Design, Synthesis, and Characterization of Ogerin-Based Positive Allosteric Modulators for G Protein-Coupled Receptor 68 (GPR68). JOURNAL OF MEDICINAL CHEMISTRY, 2019. 62(16): p. 7557–7574. Cerutis, D.R., et al., Lysophosphatidic acid regulates SPHK1 and GPR68/OGR1 in human gingival fibroblasts. FASEB JOURNAL, 2013. 27. de Valliere, C., et al., The Gene Encoding the G Protein-Coupled Receptor 68 (GPR68/OGR1) is Regulated by TNF, Hypoxia, and Low pH in Human Monocytic Cells. GASTROENTEROLOGY, 2012. 1421(5): p. S870-S870. Jiang, Z., et al., Shear stress regulates the SNAP23-mediated endothelial secretion of VWF through the GPR68/PKA/vimentin mechanotransduction pathway. BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 2022. 607: p. 166–173. Jiang, Z., et al., Shear Stress Regulates the SNAP23-mediated Endothelial secretion of VWF through the GPR68/PKA/vimentin Mechanotransduction Pathway (vol 607, pg 166, 2022). BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 2022. 609: p. 195–196. Zhao, D., et al., GPR68 Improves Nerve Damage and Myelination in an Immature Rat Model Induced by Sevoflurane Anesthesia by Activating cAMP/CREB to Mediate BDNF. ACS CHEMICAL NEUROSCIENCE, 2022. 13(3): p. 423–431. Zhou, G. and X. Zha, GPR68 Contributes to Persistent Acidosis-Induced Activation of AGC Kinases and Tyrosine Phosphorylation in Organotypic Hippocampal Slices. FRONTIERS IN NEUROSCIENCE, 2021. 15(692217). Zhou, G., T. Wang and X. Zha, RNA-Seq analysis of knocking out the neuroprotective proton-sensitive GPR68 on basal and acute ischemia-induced transcriptome changes and signaling in mouse brain. FASEB JOURNAL, 2021. 35(e214614). Huang, D.W., B.T. Sherman and R.A. Lempicki, Bioinformatics enrichment tools: paths toward the comprehensive functional analysis of large gene lists. Nucleic Acids Res, 2009. 37(1): p. 1–13. Huang, D.W., B.T. Sherman and R.A. Lempicki, Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. NATURE PROTOCOLS, 2009. 4(1): p. 44–57. Wang, L., et al., Adenosylmethionine Decarboxylase 1 (AMD1)-Mediated mRNA Processing and Cell Adhesion Activated & Inhibited Transition Mechanisms by Different Comparisons Between Chimpanzee and Human Left Hemisphere. CELL BIOCHEMISTRY AND BIOPHYSICS, 2014. 70(1): p. 279–288. Sun, L., et al., Glycogen Debranching Enzyme 6 (AGL), Enolase 1 (ENOSF1), Ectonucleotide Pyrophosphatase 2 (ENPP2_1), Glutathione S-Transferase 3 (GSTM3_3) and Mannosidase (MAN2B2) Metabolism Computational Network Analysis Between Chimpanzee and Human Left Cerebrum. CELL BIOCHEMISTRY AND BIOPHYSICS, 2011. 61(3): p. 493–505. Lin, H., et al., P-glycoprotein (ABCB1) inhibited network of mitochondrion transport along microtubule and BMP signal-induced cell shape in chimpanzee left cerebrum by systems-theoretical analysis. CELL BIOCHEMISTRY AND FUNCTION, 2012. 30(7): p. 582–587. Wang, L., et al., Signal Transducer and Activator of Transcription 2 (STAT2) Metabolism Coupling Postmitotic Outgrowth to Visual and Sound Perception Network in Human Left Cerebrum by Biocomputation. JOURNAL OF MOLECULAR NEUROSCIENCE, 2012. 47(3): p. 649–658. Caceres, M., et al., Elevated gene expression levels distinguish human from non-human primate brains. Proc Natl Acad Sci U S A, 2003. 100(22): p. 13030–5. Tusher, V.G., R. Tibshirani and G. Chu, Significance analysis of microarrays applied to the ionizing radiation response. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2001. 98(9): p. 5116–5121. Wang, Y., et al., Inferring gene regulatory networks from multiple microarray datasets. Bioinformatics, 2006. 22(19): p. 2413–20. Ma, Z., et al., Lgr5-mediated p53 Repression through PDCD5 leads to doxorubicin resistance in Hepatocellular Carcinoma. THERANOSTICS, 2019. 9(10): p. 2967–2983. Azad, I., et al., Exploring the novel heterocyclic derivatives as lead molecules for design and development of potent anticancer agents. JOURNAL OF MOLECULAR GRAPHICS & MODELLING, 2018. 81: p. 211–228. Kopalli, S.R., et al., Pectinase-treated Panax ginseng ameliorates hydrogen peroxide-induced oxidative stress in GC-2 sperm cells and modulates testicular gene expression in aged rats. JOURNAL OF GINSENG RESEARCH, 2016. 40(2): p. 185–195. Kan, W., et al., M3 Muscarinic Receptor Interaction with Phospholipase C beta(3) Determines Its Signaling Efficiency. JOURNAL OF BIOLOGICAL CHEMISTRY, 2014. 289(16): p. 11206–11218. Kugler, K.G., et al., Quantitative trait loci-dependent analysis of a gene co-expression network associated with Fusarium head blight resistance in bread wheat (Triticum aestivum L.). BMC GENOMICS, 2013. 14(728). Ji, Y., et al., Pharmacogenomics of selective serotonin reuptake inhibitor treatment for major depressive disorder: genome-wide associations and functional genomics. PHARMACOGENOMICS JOURNAL, 2013. 13(5): p. 456–463. Tables Tables 1 to 2 are available in the Supplementary Files section Additional Declarations No competing interests reported. Supplementary Files Tables.docx Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-2906322","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Short Report","associatedPublications":[],"authors":[{"id":198285102,"identity":"6f1da330-df71-464f-ad58-60519b5334fb","order_by":0,"name":"Lin Wang","email":"data:image/png;base64,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","orcid":"","institution":"Beijing University of Posts and Telecommunications","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Lin","middleName":"","lastName":"Wang","suffix":""},{"id":198285103,"identity":"eeede670-a856-46ce-99ae-633fa3b256a1","order_by":1,"name":"tao hong","email":"","orcid":"","institution":"Beijing University of Posts and Telecommunications","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"tao","middleName":"","lastName":"hong","suffix":""},{"id":198285104,"identity":"063064e4-4d93-47ce-8e0f-bab5f8ba29bf","order_by":2,"name":"Wenbin Cui","email":"","orcid":"","institution":"Beijing University of Posts and Telecommunications","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Wenbin","middleName":"","lastName":"Cui","suffix":""},{"id":198285105,"identity":"6df3f2c2-b003-47f5-af9b-1c37c042c6e7","order_by":3,"name":"Minghu Jiang","email":"","orcid":"","institution":"Tsinghua University","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Minghu","middleName":"","lastName":"Jiang","suffix":""}],"badges":[],"createdAt":"2023-05-08 06:59:30","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-2906322/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-2906322/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":36815144,"identity":"3eafbfe5-3096-4959-a8f9-b3434f083fbb","added_by":"auto","created_at":"2023-05-11 13:58:00","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":255508,"visible":true,"origin":"","legend":"\u003cp\u003eGPR68-inhibited and -activated molecular network in Thalamus by GRNInfer\u003c/p\u003e","description":"","filename":"Onlinefloatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-2906322/v1/f1d4990b62adfafb03770349.png"},{"id":36816232,"identity":"5b429b23-6368-4176-888b-d169451aec9f","added_by":"auto","created_at":"2023-05-11 14:06:00","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":63607,"visible":true,"origin":"","legend":"\u003cp\u003eGPR68-inhibited and -activated major molecular model Thalamus by GRNInfer\u003c/p\u003e","description":"","filename":"Onlinefloatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-2906322/v1/6b442801e0453899eaec0449.png"},{"id":36993895,"identity":"330a6d46-3e15-40e6-a5df-85e63578aef9","added_by":"auto","created_at":"2023-05-13 20:59:34","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":952254,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-2906322/v1/7ce1cef1-7a8e-4913-b8b4-0c64c164d39b.pdf"},{"id":36815142,"identity":"e21e4935-234b-4366-a328-730214446cfc","added_by":"auto","created_at":"2023-05-11 13:58:00","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":69085,"visible":true,"origin":"","legend":"","description":"","filename":"Tables.docx","url":"https://assets-eu.researchsquare.com/files/rs-2906322/v1/54f98be0ce160d927ef9ecab.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise based on Thalamus big data","fulltext":[{"header":"Introduction","content":"\u003cp\u003eGPR68 is widely distributed in the human body and participates in various physiological and pathological processes. Such as, GPR68 is also expressed in the nervous system, participating in a variety of physiological and pathological processes [\u003cspan additionalcitationids=\"CR2 CR3\" citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. The abnormal expression of GPR68 is related to the occurrence and development of multiple cancer [\u003cspan additionalcitationids=\"CR6 CR7 CR8 CR9 CR10 CR11 CR12\" citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. The current research mainly focuses on its regulatory mechanism[\u003cspan additionalcitationids=\"CR15 CR16 CR17\" citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e], signaling pathways[\u003cspan additionalcitationids=\"CR20 CR21 CR22\" citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e] and potential clinical applications.\u003c/p\u003e \u003cp\u003eGPR68 locates plasma membrane, integral component of plasma membrane, functions as protein coupled receptor activity, inflammatory response, G protein coupled receptor signaling pathway, positive regulation of insulin secretion involved in cellular response to glucose stimulus, negative regulation of monocyte differentiation, cellular response to pH, positive regulation of osteoclast development, GPCRDB class A rhodopsin like, rhodopsin like receptor activity, osteoclasts by the Database for Annotation, Visualization and Integrated Discovery (DAVID) [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. However, GPR68-inhibited nucleus or ER to exosome small molecule-based circadian exercise mechanism is not clear in Thalamus.\u003c/p\u003e \u003cp\u003eWe have already published several papers[\u003cspan additionalcitationids=\"CR27 CR28\" citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e] from GDS2678[\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e] by integrating microarray significance analysis (SAM) [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e], SPSS correlation coefficient Pearson and gene (protein) reconstruction network (GRNInfer) [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e] mutual positive verification. GPR68-inhibited subnetwork we identified includes upstream ALG8, MAF, feedback SMA4, downstream GSTM3_1, GSTM3_2, NUP214, TYMS, UBB. GPR68-activated subnetwork includes upstream PER2, RAD50, feedback SPAG9, downstream CDR2, CGRRF1, DZIP3, ISCA1, RFK in GDS2678 in Thalamus.\u003c/p\u003e \u003cp\u003eThe main GPR68-inhibited and -activated molecular network will be constructed up/downstream/feedback in Thalamus for inhibited nucleus or ER to exosome non-protein based circadian exercise based on richness. The inhibition relationship in Thalamus for negative main molecular verification. The activation relationship will be found in Thalamus for positive main molecular verification.\u003c/p\u003e \u003cp\u003eThe main knowledge of GPR68-inhibited and -activated subnetwork will be constructed up/downstream/feedback for inhibited nucleus or ER to exosome non-protein based circadian exercise. We will integrate GPR68-inhibited and -activated knowledge and put forwards Thalamus model.\u003c/p\u003e \u003cp\u003eGPR68-inhibited and -activated knowledge subnetwork will be demonstrated in Thalamus for mutual negative knowledge verification. and will be positive verified by the other similar and common inhibited and/or activated knowledge.\u003c/p\u003e \u003cp\u003eThe role and mechanism of our GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise contributes to better detection, evaluation, intervention, tracking, and other health management of brain diseases.\u003c/p\u003e"},{"header":"Method","content":"\u003cp\u003e441 significant high expression molecules in 14 human left hemispheres were selected from 12,558 genes compared with the corresponding low expression of 15 chimpanzee left hemispheres in GDS2678 (public free from NCBI) by significance analysis of microarrays (SAM) (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://www-stat.stanford.edu/~tibs/SAM/\u003c/span\u003e\u003cspan address=\"http://www-stat.stanford.edu/~tibs/SAM/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e), Data were processed using a log base of two and two unpaired classes with minimum fold change (\u0026ge;\u0026thinsp;2). A false-discovery rate of 0% was chosen.\u003c/p\u003e \u003cp\u003eGPR68-inhibited molecules in Thalamus was identified negative correlation coefficient (CC \u0026le; -0.25), including ALG8, MAF, SMA4, GSTM3_1, GSTM3_2, NUP214, TYMS, UBB, which are mutual positive Pearson correlation (CC\u0026thinsp;\u0026ge;\u0026thinsp;0.25) molecules except GPR68 by SPSS.\u003c/p\u003e \u003cp\u003eGPR68 activated molecules from high expression Pearson mutual positive correlation coefficient (CC\u0026thinsp;\u0026ge;\u0026thinsp;0.25) were selected including PER2, RAD50, SPAG9, CDR2, CGRRF1, DZIP3, ISCA1, RFK in Thalamus by SPSS.\u003c/p\u003e \u003cp\u003eGRNInfer is a tool used to construct all molecular activation and inhibition direction total network by linear programming and decomposition procedure defined by the following equation:\u003cdiv id=\"Equ1\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equ1\" name=\"EquationSource\"\u003e\n$$\\mathcal{J}=\\left(\\dot{\\text{X}}-\\text{B}\\right)\\text{U}{\\text{E}}^{-1}{\\text{V}}^{\\text{T}}+\\text{Y}{\\text{V}}^{\\text{T}}=\\widehat{\\mathcal{I}}+\\text{Y}{\\text{V}}^{\\text{T}}$$\u003c/div\u003e\u003cdiv class=\"EquationNumber\"\u003e1\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eThe main GPR68-inhibited and -activated molecular network was constructed up/downstream/feedback in Thalamus for inhibited nucleus or ER to exosome non-protein based circadian exercise by GRNInfer based on richness.\u003c/p\u003e \u003cp\u003eThe inhibition relationship was found in Thalamus for negative main molecular verification. The activation relationship in Thalamus for positive main molecular verification.\u003c/p\u003e \u003cp\u003eThe main knowledge of GPR68-inhibited and activated subnetwork was constructed up/downstream/feedback for inhibited nucleus or ER to exosome non-protein based circadian exercise in Thalamus by DAVID, respectively, including GOTERM-BP-DIRECT, GOTERM-MF-DIRECT, KEGG-PATHWAY, BIOCARTA, GenMAPP, GOTERM-CC-DIRECT, GNF-U133A-QUARTILE UNIGENE-EST-QUARTILE.\u003c/p\u003e \u003cp\u003eWe integrated GPR68-inhibited and -activated knowledge and put forwards our model. GPR68-inhibited and -activated knowledge subnetwork was setup in Thalamus for mutual negative knowledge verification. Our model was positive verified by the corresponding other similar and common inhibited and/or activated knowledge.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eGPR68-inhibited and -activated network was constructed in Thalamus. GPR68 inhibited molecular subnetwork composed of upstream ALG8, MAF, feedback SMA4, and downstream GSTM3_1, GSTM3_2, NUP214, TYMS, UBB in Thalamus (Figure 1). GPR68 activated subnetwork included upstream PER2, RAD50, feedback SPAG9, and downstream CDR2, CGRRF1, DZIP3, ISCA1, RFK in Thalamus (Figure 1), based on mutual positive verification using CC and GRNInfer.\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;The main GPR68 inhibited molecular subnetwork composed of upstream ALG8, or MAF, feedback SMA4, and downstream GSTM3_1 with TYMS (Figure 2). GPR68 activated molecular subnetwork contained upstream RAD50, feedback SPAG9, and downstream RFK (Figure 2).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe relationship of SMA4 inhibition to SPAG9 -\u0026gt; MAF -\u0026gt; RAD50 -\u0026gt; ALG8 was found in Thalamus The relationship of ALG8 activation to SMA4 -\u0026gt; GSTM3_1 -\u0026gt; TYMS -\u0026gt; MAF in Thalamus (Figure 2).\u003c/p\u003e\n\u003cp\u003eThe main GPR68 inhibited knowledge subnetwork was identified upstream nucleus sequence specific DNA binding, or ER amino acid glycosylation, feedback nucleoplasm RNA splicing, and downstream extracellular exosome small molecule-based circadian exercise in Thalamus by DAVID, as shown in Table 1. The main GPR68 activated subnetwork included upstream membrane positive regulation of kinase activity, feedback integral component of membrane kinesin binding, and downstream mitochondrion apoptotic process in Thalamus by DAVID，as shown in Table 2.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe main GPR68 inhibited common knowledge appears nucleus appears in the subnetwork of GPR68 inhibited-upstream MAF and inhibited-downstream GSTM3_1, TYMS in Thalamus. The main GPR68 activated common knowledge appears metal ion binding appears in the subnetwork of GPR68 activated-upstream RAD50 and activated-downstream RFK; ATP binding in the subnetwork of GPR68 activated-upstream RAD50 and activated-downstream RFK in Thalamus.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe main GPR68 inhibited and activated common knowledge appears small molecule metabolic process appears in the subnetwork of GPR68 inhibited-downstream GSTM3_1, TYMS and activated-downstream RFK; protein binding in the subnetwork of GPR68 inhibited-upstream MAF and activated-upstream RAD50 and activated-feedback SPAG9; nucleoplasm in the subnetwork of GPR68 inhibited-feedback SMA4 and inhibited-downstream TYMS and activated-upstream RAD50; mitochondrion in the subnetwork of GPR68 inhibited-downstream TYMS and activated-downstream RFK; Metabolic pathways in the subnetwork of GPR68 inhibited-upstream ALG8 and inhibited-downstream TYMS and activated-downstream RFK; integral component of membrane in the subnetwork of GPR68 inhibited-upstream ALG8 and activated-feedback SPAG9; extracellular exosome in the subnetwork of GPR68 inhibited-downstream GSTM3_1 and activated-feedback SPAG9; cytosol in the subnetwork of GPR68 inhibited-downstream GSTM3_1, TYMS and activated-feedback SPAG9 and activated-downstream RFK; cytoplasm in the subnetwork of GPR68 inhibited-upstream MAF and inhibited-downstream GSTM3_1, TYMS and activated-feedback SPAG9 and activated-downstream RFK in Thalamus, as shown in Table 1 and 2.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eGPR68-inhibited and -activated network was constructed in Thalamus. GPR68 inhibited molecular subnetwork composed of upstream ALG8, MAF, feedback SMA4, and downstream GSTM3_1, GSTM3_2, NUP214, TYMS, UBB in Thalamus (Figure 1). GPR68 activated subnetwork included upstream PER2, RAD50, feedback SPAG9, and downstream CDR2, CGRRF1, DZIP3, ISCA1, RFK in Thalamus (Figure 1), based on mutual positive verification using CC and GRNInfer.\u003c/p\u003e\n\u003cp\u003eThe main GPR68 activated molecular subnetwork composed of upstream ALG8, or MAF, feedback SMA4, and downstream GSTM3_1 with TYMS. GPR68 inhibited molecular subnetwork contained upstream RAD50, feedback SPAG9, and downstream RFK (Figure 2).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe relationship of SMA4 inhibition to SPAG9 -\u0026gt; MAF -\u0026gt; RAD50 -\u0026gt; ALG8 was found in Thalamus for negative main molecular verification. The relationship of ALG8 activation to SMA4 -\u0026gt; GSTM3_1 -\u0026gt; TYMS -\u0026gt; MAF in Thalamus for positive main molecular verification (Figure 2).\u003c/p\u003e\n\u003cp\u003eThe main GPR68 inhibited upstream molecule was determined as ALG8, or MAF in Thalamus by GRNInfer (Figure 1-2). The corresponding main inhibited upstream knowledge subnetwork was identified as nucleus sequence specific DNA binding, or ER amino acid glycosylation based on DAVID. We proposed GPR68 inhibited upstream nucleus sequence specific DNA binding, or ER amino acid glycosylation in Thalamus.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eALG8 refers to ALG8 alpha-1,3-glucosyltransferase, an enzyme in the endoplasmic reticulum membrane and an integral component of the membrane. It has alpha 1,3 mannosyltransferase activity and dolichyl pyrophosphate Man9GlcNAc2 alpha 1,3 glucosyltransferase activity. ALG8 is involved in various biological processes, including protein N-linked glycosylation, dolichol-linked oligosaccharide biosynthetic process, oligosaccharide lipid intermediate biosynthetic process, protein N-linked glycosylation via asparagine, post-translational protein modification, cellular protein metabolic process, and mannosylation. ALG8 is associated with N-glycan biosynthesis and metabolic pathways. It is also related to glycoprotein metabolism, protein amino acid glycosylation, glycoprotein biosynthesis, and biopolymer glycosylation.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eMAF (v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog) is a protein present in chromatin, nucleus, and cytoplasm. it possesses transcriptional activator activity and sequence-specific DNA binding and protein binding capabilities. MAF is involved in various biological processes, such as negative regulation of transcription from RNA polymerase II promoter, cytokine production, regulation of transcription from RNA polymerase II promoter, transcription from RNA polymerase II promoter, regulation of chondrocyte differentiation, positive regulation of transcription from RNA polymerase II promoter, cell development, inner ear development, and lens fiber cell differentiation. It is associated with transcriptional misregulation in cancer and inflammatory bowel disease (IBD). MAF participates in activating the Th2 cytokine genes expression and has effects on RNA polymerase II transcription factor activity was selected by DAVID (Table 1) as positive verification for the main Thalamus knowledge.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe main GPR68 inhibited feedback molecule was determined as SMA4 in Thalamus by GRNInfer (Figure 1-2). The corresponding main inhibited feedback knowledge subnetwork was identified as nucleoplasm RNA splicing based on DAVID. We proposed GPR68 inhibited feedback nucleoplasm RNA splicing in Thalamus. SMA4 refers to the glucuronidase beta pseudogene (SMA4), which can be found in the nucleoplasm, protein complex assembly, nuclear mRNA splicing via spliceosome, RNA splicing, RNA splicing via transesterification reactions with bulged adenosine as nucleophile, RNA splicing via transesterification reactions, and mRNA processing was selected by DAVID (Table 1) as positive verification for the main Thalamus knowledge.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe main GPR68 inhibited downstream molecule was determined as GSTM3_1 with TYMS in Thalamus by GRNInfer (Figure 1-2). The corresponding main inhibited downstream knowledge subnetwork was identified as extracellular exosome small molecule-based circadian exercise based on DAVID. We proposed GPR68 inhibited downstream extracellular exosome small molecule-based circadian exercise in Thalamus.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eGSTM3_1 refers to glutathione S-transferase mu 3 (brain) (GSTM3), which can be found in the nucleus, cytoplasm, cytosol, sperm fibrous sheath, and extracellular exosome. It has glutathione transferase activity, enzyme binding ability, identical protein binding ability, protein homodimerization activity, and glutathione binding ability. GSTM3_1 is involved in various biological processes, such as glutathione metabolic process, xenobiotic metabolic process, establishment of blood nerve barrier, nitrobenzene metabolic process, xenobiotic catabolic process, response to estrogen, small molecule metabolic process, cellular detoxification of nitrogen compounds, and glutathione derivative biosynthetic process. GSTM3_1 is related to glutathione metabolism, metabolism of xenobiotics by cytochrome P450, drug metabolism cytochrome P450, and chemical carcinogenesis. Additionally, it has an association with circadian exercise.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eTYMS (Thymidylate Synthetase) is an enzyme that is present in the nucleus, nucleoplasm, nucleolus, cytoplasm, mitochondrion, mitochondrial inner membrane, mitochondrial matrix, and cytosol. It possesses nucleotide binding, mRNA binding, thymidylate synthase activity, folic acid binding, drug binding, protein homodimerization activity, and cofactor binding. TYMS is involved in various biological processes, including G1/S transition of mitotic cell cycle, regulation of transcription involved in G1/S transition of mitotic cell cycle, mitotic cell cycle, nucleobase containing compound metabolic process, pyrimidine nucleobase metabolic process, dTMP biosynthetic process, dTTP biosynthetic process, aging, circadian rhythm, cell proliferation, deoxyribonucleoside monophosphate biosynthetic process, response to toxic substance, cell growth, immortalization of host cell by virus, uracil metabolic process, organ regeneration, methylation, response to progesterone, response to vitamin A, response to cytokine, tetrahydrofolate interconversion, response to drug, small molecule metabolic process, response to ethanol, dUMP metabolic process, pyrimidine nucleoside biosynthetic process, response to organophosphorus, developmental growth, cartilage development, response to glucocorticoid, response to folic acid, nucleobase containing small molecule metabolic process, intestinal epithelial cell maturation, and DNA biosynthetic process. TYMS is involved in metabolic pathways such as Pyrimidine metabolism and One carbon pool by folate was selected by DAVID (Table 1) as positive verification for the main Thalamus knowledge. There are literature supporting the relationship between GPR68 and RAD50 families\u0026nbsp;[33-36].\u003c/p\u003e\n\u003cp\u003eThe main GPR68 activated upstream molecule was determined as RAD50 in Thalamus by GRNInfer (Figure 1-2). The corresponding main activated upstream knowledge subnetwork was identified as membrane positive regulation of kinase activity based on DAVID. We proposed GPR68 activated upstream membrane positive regulation of kinase activity in Thalamus. RAD50 refers to RAD50 double-strand break repair protein (RAD50), which can be found in the nuclear chromosome telomeric region, nucleoplasm, membrane, Mre11 complex, site of double-strand break, and pronucleus. RAD50 has single-stranded DNA endodeoxyribonuclease activity, DNA binding ability, ATP-dependent DNA helicase activity, protein binding ability, ATP binding ability, 3\u0026rsquo; 5\u0026rsquo; exonuclease activity, protein binding bridging ability, and metal ion binding ability. RAD50 is involved in the regulation of mitotic recombination, telomere maintenance, double-strand break repair via homologous recombination, DNA repair, double-strand break repair, double-strand break repair via nonhomologous end joining, DNA recombination, cellular response to DNA damage stimulus, telomere maintenance via telomerase, reciprocal meiotic recombination, regulation of mitotic cell cycle, viral process, telomeric 3\u0026rsquo; overhang formation, positive regulation of protein autophosphorylation, positive regulation of telomere maintenance, DNA duplex unwinding, positive regulation of kinase activity, double-strand break repair via synthesis-dependent strand annealing, chromosome organization involved in meiosis, nucleic acid phosphodiester bond hydrolysis, and negative regulation of telomere capping. RAD50 is related to homologous recombination and non-homologous end joining. It is also involved in the h_atmPathway: ATM Signaling Pathway, h_atrbrcaPathway: Role of BRCA1, BRCA2 and ATR in Cancer Susceptibility. Additionally, RAD50 is related to the cell cycle, response to an endogenous stimulus, M phase, and response to DNA damage stimulus was selected by DAVID (Table 2) as positive verification for the main Thalamus knowledge.\u003c/p\u003e\n\u003cp\u003eThe main GPR68 activated feedback molecule was determined as SPAG9 in Thalamus by GRNInfer (Figure 1-2). The corresponding main activated feedback knowledge subnetwork was identified as integral component of membrane kinesin binding based on DAVID. We proposed GPR68 activated feedback integral component of membrane kinesin binding in Thalamus. SPAG9, which stands for sperm associated antigen 9, is a protein that is associated with various cellular components such as acrosomal vesicles, cytoplasm, microtubule organizing centers, cytosol, integral components of membranes, perinuclear regions of cytoplasm, and extracellular exosomes. It exhibits multiple molecular functions such as MAP kinase scaffold activity, protein binding, JUN kinase binding, kinesin binding, receptor signaling complex scaffold activity, and mitogen activated protein kinase p38 binding. Its biological processes include the activation of JUN kinase activity, spermatogenesis, positive regulation of cell migration, retrograde transport endosome to Golgi, muscle cell differentiation, positive regulation of neuron differentiation, striated muscle cell differentiation, positive regulation of muscle cell differentiation, protein homooligomerization, and negative regulation of protein homodimerization activity was selected by DAVID (Table 2) as positive verification for the main Thalamus knowledge.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe main GPR68 activated downstream molecule was determined as RFK in Thalamus by GRNInfer (Figure 1-2). The corresponding main activated downstream knowledge subnetwork was identified as mitochondrion apoptotic process based on DAVID. We proposed GPR68 activated downstream mitochondrion apoptotic process in Thalamus. Riboflavin kinase (RFK) is a protein that is located in the cytoplasm, mitochondria, and cytosol. It exhibits molecular functions such as ATP binding, riboflavin kinase activity, and metal ion binding. RFK is involved in vitamin metabolic processes, including water-soluble vitamin metabolic processes, riboflavin metabolic processes, and riboflavin biosynthetic processes. It also participates in apoptotic processes, FMN biosynthetic processes, phosphorylation, positive regulation of NAD\u0026sect;H oxidase activity, small molecule metabolic processes, and reactive oxygen species metabolic processes. The functions of RFK are usually associated with riboflavin metabolism and metabolic pathways. Its important molecular function is magnesium ion binding was selected by DAVID (Table 2) as positive verification for the main Thalamus knowledge. There are literature supporting the activated relationship between GPR68 and RFK families[37, 38].\u003c/p\u003e\n\u003cp\u003eWe put forwards the model of GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise based on Thalamus big data in Thalamus. GPR68 inhibited nucleoplasm RNA splicing coupling nucleus sequence specific DNA binding, or ER amino acid glycosylation to extracellular exosome small molecule-based circadian exercise, GPR68 activated integral component of membrane kinesin binding coupling membrane positive regulation of kinase activity to mitochondrion apoptotic process for mutual negative knowledge verification.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe main GPR68-inhibited model is positive verified by common knowledge，including nucleus appears in the subnetwork of GPR68 inhibited-upstream MAF and inhibited-downstream GSTM3_1, TYMS in Thalamus. The main GPR68-activated model is positive verified by the common knowledge, including metal ion binding appears in the subnetwork of GPR68 activated-upstream RAD50 and activated-downstream RFK; ATP binding in the subnetwork of GPR68 activated-upstream RAD50 and activated-downstream RFK in Thalamus.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe main GPR68 inhibited and activated model was positive verified by common knowledge, containing small molecule metabolic process appears in the subnetwork of GPR68 inhibited-downstream GSTM3_1, TYMS and activated-downstream RFK; protein binding in the subnetwork of GPR68 inhibited-upstream MAF and activated-upstream RAD50 and activated-feedback SPAG9; nucleoplasm in the subnetwork of GPR68 inhibited-feedback SMA4 and inhibited-downstream TYMS and activated-upstream RAD50; mitochondrion in the subnetwork of GPR68 inhibited-downstream TYMS and activated-downstream RFK; Metabolic pathways in the subnetwork of GPR68 inhibited-upstream ALG8 and inhibited-downstream TYMS and activated-downstream RFK; integral component of membrane in the subnetwork of GPR68 inhibited-upstream ALG8 and activated-feedback SPAG9; extracellular exosome in the subnetwork of GPR68 inhibited-downstream GSTM3_1 and activated-feedback SPAG9; cytosol in the subnetwork of GPR68 inhibited-downstream GSTM3_1, TYMS and activated-feedback SPAG9 and activated-downstream RFK; cytoplasm in the subnetwork of GPR68 inhibited-upstream MAF and inhibited-downstream GSTM3_1, TYMS and activated-feedback SPAG9 and activated-downstream RFK in Thalamus (Table 1 and 2).\u003c/p\u003e"},{"header":"Summary","content":"\u003cp\u003eThe inhibition relationships among the main molecules were found in Thalamus for negative molecular verification. The activation relationships among the main molecules were found in Thalamus for positive molecular verification.\u003c/p\u003e \u003cp\u003eWe put forwards GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise in Thalamus, based on GPR68 inhibited nucleoplasm RNA splicing coupling nucleus sequence specific DNA binding, or ER amino acid glycosylation to extracellular exosome small molecule-based circadian exercise, and GPR68 activated integral component of membrane kinesin binding coupling membrane positive regulation of kinase activity to mitochondrion apoptotic process for mutual negative knowledge verification. Our model is positive verified by the other similar and common inhibited and/or activated knowledge in Thalamus.\u003c/p\u003e \u003cp\u003eThe role and mechanism of our GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise contributes to better detection, evaluation, intervention, tracking, and other health management of brain diseases.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eEthics approval and consent to participate\u003c/h2\u003e\n\u003cp\u003eWe have not Ethics because our data GSE10140-10141 public free from NCBI.\u003c/p\u003e\n\u003ch2\u003eConsent for publication\u003c/h2\u003e\n\u003cp\u003eAll authors have approved the manuscript for submission.\u003c/p\u003e\n\u003ch2\u003eData Availability Statement\u003c/h2\u003e\n\u003cp\u003eOur data GSE10140-10141 public free from NCBI.\u003c/p\u003e\n\u003ch2\u003eCompeting interests\u0026nbsp;\u003c/h2\u003e\n\u003cp\u003eWe have not any competing interests with others.\u003c/p\u003e\n\u003ch2\u003eFunding\u0026nbsp;\u003c/h2\u003e\n\u003cp\u003eThis work is supported by grants from the National Natural Science Key Foundation of China (62036001), National Social Science Major Foundation of China (14ZDB154 * 15ZDB017), and the Independent scientific research project of Tsinghua University (20161080056).\u003c/p\u003e\n\u003ch2\u003eAuthor Contribution Statement\u003c/h2\u003e\n\u003cp\u003eLW not only has designed the whole experiment, put forward hypotheses and written the paper, but also established SAM and GRNInfer databases. TH has constructed molecular and knowledge network，Pearson positive correlation coefficient (CC) database. WBC has done programme and setup DAVID database. MHJ has done figures and tables and help analysis in this paper. All authors read and approved the final Manuscript.\u003c/p\u003e\n\u003ch2\u003eAcknowledgments\u003c/h2\u003e\n\u003cp\u003eThank Zhenfu Jiang from Tencent for suggesting us many tools and methods.\u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eWang, T., et al., GPR68 Is a Neuroprotective Proton Receptor in Brain Ischemia. STROKE, 2020. 51(12): p.\u0026nbsp;3690\u0026ndash;3700.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eXu, Y., M.T. Lin and X. Zha, GPR68 deletion impairs hippocampal long-term potentiation and passive avoidance behavior (vol\u0026nbsp;13, pg 132, 2020). MOLECULAR BRAIN, 2021. 14(401).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eXu, Y., M.T. Lin and X. Zha, GPR68 deletion impairs hippocampal long-term potentiation and passive avoidance behavior. MOLECULAR BRAIN, 2020. 13(1321).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNogueira, P.A.S., et al., The orphan receptor GPR68 is expressed in the hypothalamus and is involved in the regulation of feeding. NEUROSCIENCE LETTERS, 2022. 781(136660).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWiley, S.Z., et al., GPR68: An Emerging Drug Target in Cancer. INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES, 2019. 20(5593).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShore, D., et al., GPR68 limits the severity of chemical-induced oral epithelial dysplasia. Scientific reports, 2023. 13(1): p.\u0026nbsp;353\u0026ndash;353.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHe, X., et al., Proton Sensor GPR68 Is Essential to Maintain Myeloid Malignancies. BLOOD, 2018. 1321.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eElemam, N.M., et al., Expression Status of Proton Sensing Receptor GPR68 in Various Breast Cancer Molecular Subtypes. FASEB JOURNAL, 2022. 361.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eElemam, N.M., et al., Expression of GPR68, an Acid-Sensing Orphan G Protein-Coupled Receptor, in Breast Cancer. FRONTIERS IN ONCOLOGY, 2022. 12(847543).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang, W., et al., Clinical data analysis reveals the role of OGR1 (GPR68) in head and neck squamous cancer. Animal models and experimental medicine, 2020. 3(1): p.\u0026nbsp;55\u0026ndash;61.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWiley, S.Z., et al., GPR68, a proton-sensing GPCR that mediates interaction of pancreatic cancer associated fibroblasts and cancer cells, is a potential therapeutic target for pancreatic cancer. CANCER RESEARCH, 2019. 79S(13).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWiley, S., et al., GPR68, a proton sensing GPCR, mediates interaction of pancreatic cancer associated fibroblasts and cancer cells. FASEB JOURNAL, 2018. 32S(1).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWiley, S.Z., et al., GPR68, a proton-sensing GPCR, mediates interaction of cancer-associated fibroblasts and cancer cells. FASEB JOURNAL, 2018. 32(3): p.\u0026nbsp;1170\u0026ndash;1183.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHuang, X., et al., Allosteric ligands for the pharmacologically dark receptors GPR68 and GPR65. NATURE, 2015. 527(7579): p.\u0026nbsp;477-+.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBell, T.J., et al., Inhibition of Myofibroblast Differentiation and SMAD Mediated Transcription via Small Molecule Stimulation of GPR68. AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE, 2021. 203S(9).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYu, X., et al., Design, Synthesis, and Characterization of Ogerin-Based Positive Allosteric Modulators for G Protein-Coupled Receptor 68 (GPR68). JOURNAL OF MEDICINAL CHEMISTRY, 2019. 62(16): p.\u0026nbsp;7557\u0026ndash;7574.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCerutis, D.R., et al., Lysophosphatidic acid regulates SPHK1 and GPR68/OGR1 in human gingival fibroblasts. FASEB JOURNAL, 2013. 27.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ede Valliere, C., et al., The Gene Encoding the G Protein-Coupled Receptor 68 (GPR68/OGR1) is Regulated by TNF, Hypoxia, and Low pH in Human Monocytic Cells. GASTROENTEROLOGY, 2012. 1421(5): p.\u0026nbsp;S870-S870.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJiang, Z., et al., Shear stress regulates the SNAP23-mediated endothelial secretion of VWF through the GPR68/PKA/vimentin mechanotransduction pathway. BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 2022. 607: p.\u0026nbsp;166\u0026ndash;173.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJiang, Z., et al., Shear Stress Regulates the SNAP23-mediated Endothelial secretion of VWF through the GPR68/PKA/vimentin Mechanotransduction Pathway (vol\u0026nbsp;607, pg 166, 2022). BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 2022. 609: p.\u0026nbsp;195\u0026ndash;196.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhao, D., et al., GPR68 Improves Nerve Damage and Myelination in an Immature Rat Model Induced by Sevoflurane Anesthesia by Activating cAMP/CREB to Mediate BDNF. ACS CHEMICAL NEUROSCIENCE, 2022. 13(3): p.\u0026nbsp;423\u0026ndash;431.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhou, G. and X. Zha, GPR68 Contributes to Persistent Acidosis-Induced Activation of AGC Kinases and Tyrosine Phosphorylation in Organotypic Hippocampal Slices. FRONTIERS IN NEUROSCIENCE, 2021. 15(692217).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhou, G., T. Wang and X. Zha, RNA-Seq analysis of knocking out the neuroprotective proton-sensitive GPR68 on basal and acute ischemia-induced transcriptome changes and signaling in mouse brain. FASEB JOURNAL, 2021. 35(e214614).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHuang, D.W., B.T. Sherman and R.A. Lempicki, Bioinformatics enrichment tools: paths toward the comprehensive functional analysis of large gene lists. Nucleic Acids Res, 2009. 37(1): p.\u0026nbsp;1\u0026ndash;13.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHuang, D.W., B.T. Sherman and R.A. Lempicki, Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. NATURE PROTOCOLS, 2009. 4(1): p.\u0026nbsp;44\u0026ndash;57.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang, L., et al., Adenosylmethionine Decarboxylase 1 (AMD1)-Mediated mRNA Processing and Cell Adhesion Activated \u0026amp; Inhibited Transition Mechanisms by Different Comparisons Between Chimpanzee and Human Left Hemisphere. CELL BIOCHEMISTRY AND BIOPHYSICS, 2014. 70(1): p.\u0026nbsp;279\u0026ndash;288.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSun, L., et al., Glycogen Debranching Enzyme 6 (AGL), Enolase 1 (ENOSF1), Ectonucleotide Pyrophosphatase 2 (ENPP2_1), Glutathione S-Transferase 3 (GSTM3_3) and Mannosidase (MAN2B2) Metabolism Computational Network Analysis Between Chimpanzee and Human Left Cerebrum. CELL BIOCHEMISTRY AND BIOPHYSICS, 2011. 61(3): p.\u0026nbsp;493\u0026ndash;505.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLin, H., et al., P-glycoprotein (ABCB1) inhibited network of mitochondrion transport along microtubule and BMP signal-induced cell shape in chimpanzee left cerebrum by systems-theoretical analysis. CELL BIOCHEMISTRY AND FUNCTION, 2012. 30(7): p.\u0026nbsp;582\u0026ndash;587.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang, L., et al., Signal Transducer and Activator of Transcription 2 (STAT2) Metabolism Coupling Postmitotic Outgrowth to Visual and Sound Perception Network in Human Left Cerebrum by Biocomputation. JOURNAL OF MOLECULAR NEUROSCIENCE, 2012. 47(3): p.\u0026nbsp;649\u0026ndash;658.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCaceres, M., et al., Elevated gene expression levels distinguish human from non-human primate brains. Proc Natl Acad Sci U S A, 2003. 100(22): p.\u0026nbsp;13030\u0026ndash;5.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTusher, V.G., R. Tibshirani and G. Chu, Significance analysis of microarrays applied to the ionizing radiation response. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2001. 98(9): p.\u0026nbsp;5116\u0026ndash;5121.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang, Y., et al., Inferring gene regulatory networks from multiple microarray datasets. Bioinformatics, 2006. 22(19): p.\u0026nbsp;2413\u0026ndash;20.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMa, Z., et al., Lgr5-mediated p53 Repression through PDCD5 leads to doxorubicin resistance in Hepatocellular Carcinoma. THERANOSTICS, 2019. 9(10): p.\u0026nbsp;2967\u0026ndash;2983.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAzad, I., et al., Exploring the novel heterocyclic derivatives as lead molecules for design and development of potent anticancer agents. JOURNAL OF MOLECULAR GRAPHICS \u0026amp; MODELLING, 2018. 81: p.\u0026nbsp;211\u0026ndash;228.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKopalli, S.R., et al., Pectinase-treated Panax ginseng ameliorates hydrogen peroxide-induced oxidative stress in GC-2 sperm cells and modulates testicular gene expression in aged rats. JOURNAL OF GINSENG RESEARCH, 2016. 40(2): p.\u0026nbsp;185\u0026ndash;195.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKan, W., et al., M3 Muscarinic Receptor Interaction with Phospholipase C beta(3) Determines Its Signaling Efficiency. JOURNAL OF BIOLOGICAL CHEMISTRY, 2014. 289(16): p.\u0026nbsp;11206\u0026ndash;11218.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKugler, K.G., et al., Quantitative trait loci-dependent analysis of a gene co-expression network associated with Fusarium head blight resistance in bread wheat (Triticum aestivum L.). BMC GENOMICS, 2013. 14(728).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJi, Y., et al., Pharmacogenomics of selective serotonin reuptake inhibitor treatment for major depressive disorder: genome-wide associations and functional genomics. PHARMACOGENOMICS JOURNAL, 2013. 13(5): p.\u0026nbsp;456\u0026ndash;463.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003eTables 1 to 2 are available in the Supplementary Files section\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"GPR68 inhibited Thalamus subnetwork, GPR68 activated Thalamus subnetwork, GPR68 inhibited nucleus or ER to exosome, GPR68 inhibited circadian exercise, GPR68 activated protein","lastPublishedDoi":"10.21203/rs.3.rs-2906322/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-2906322/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eG protein-coupled receptor (GPR68) is widely distributed in the human body and participates in various physiological and pathological processes. The main GPR68-inhibited and -activated molecular network was constructed in Thalamus from GDS2678 of NCBI by integrating SAM, SPSS Pearson and GRNInfer mutual positive verification based on richness. The main GPR68 inhibited molecular subnetwork composed of upstream ALG8, or MAF, feedback SMA4, and downstream GSTM3_1 with TYMS. GPR68 activated molecular subnetwork contained upstream RAD50, feedback SPAG9, and downstream RFK. The relationship of SMA4 inhibition to SPAG9 -\u0026gt; MAF -\u0026gt; RAD50 -\u0026gt; ALG8 was found in Thalamus for negative main molecular verification. The relationship of ALG8 activation to SMA4 -\u0026gt; GSTM3_1 -\u0026gt; TYMS -\u0026gt; MAF in Thalamus for positive main molecular verification. Our results show GPR68 inhibited SMA4-nucleoplasm RNA splicing coupling ALG8, or MAF-nucleus sequence specific DNA binding, or ER amino acid glycosylation to GSTM3_1 with TYMS-extracellular exosome small molecule-based circadian exercise, and GPR68 activated SPAG9-integral component of membrane kinesin binding coupling RAD50-membrane positive regulation of kinase activity to RFK-mitochondrion apoptotic process for mutual negative knowledge verification. We put forwards GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise based on Thalamus big data. Our model is positive verified by the other similar and common inhibited and/or activated knowledge in Thalamus. The role and mechanism of our GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise contributes to better detection, evaluation, intervention, tracking, and other health management of brain diseases.\u003c/p\u003e","manuscriptTitle":"GPR68 inhibited nucleus or ER to exosome non-protein based circadian exercise based on Thalamus big data","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2023-05-11 13:57:55","doi":"10.21203/rs.3.rs-2906322/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"fa742f94-2033-4f66-9b32-10a54ef56d5a","owner":[],"postedDate":"May 11th, 2023","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2023-05-13T20:59:22+00:00","versionOfRecord":[],"versionCreatedAt":"2023-05-11 13:57:55","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-2906322","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-2906322","identity":"rs-2906322","version":["v1"]},"buildId":"ehx78VzkSd0WSzXnipQa-","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}